1376: Molecular Diagnostic Detection of Haploid Germ Cells in Testicular Biopsy Specimens by Online RT-PCR

2004 ◽  
Vol 171 (4S) ◽  
pp. 362-363
Author(s):  
Mark G. Schrader ◽  
Markus Muller ◽  
Wolfgang Schulze ◽  
Steffen Weikert ◽  
Kurt Miller
Keyword(s):  
Germ Cells ◽  
Testicular Biopsy ◽  
Molecular Diagnostic ◽  
Rt Pcr ◽  
Biopsy Specimens ◽  
Diagnostic Detection

scholarly journals Molecular Detection of Mycobacterium tuberculosis in Oral Mucosa from Patients with Presumptive Tuberculosis

2020 ◽  
Vol 9 (12) ◽  
pp. 4124
Author(s):  
Barbara Molina-Moya ◽  
Nelly Ciobanu ◽  
Marta Hernandez ◽  
Cristina Prat-Aymerich ◽  
Valeriu Crudu ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Oral Mucosa ◽  
The Elderly ◽  
Storage Conditions ◽  
Molecular Diagnostic ◽  
Smear Microscopy ◽  
X Rays ◽  
Rt Pcr ◽  
Tuberculosis Complex ◽  
Tb 34

Tuberculosis (TB) diagnosis is increasingly based on the detection of Mycobacterium tuberculosis complex (MTBC) DNA in sputum using molecular diagnostic tests as the first test for diagnosis. However, sputum can be difficult to obtain in children, patients without productive cough, and the elderly and approaches testing non-sputum samples are needed. We evaluated whether TB can be detected from the oral mucosa of patients with TB. Adults with presumptive TB were examined using culture, Xpert MTB/RIF, smear microscopy and X-Rays. Oral mucosa swabs collected on PrimeStore-MTM, stored at room temperature if tested within 30 days or at −20 °C if examined at a later time. RT-PCR was performed to detect M. tuberculosis DNA. Eighty patients had bacteriologically-confirmed TB, 34 had bacteriologically-negative TB (negative tests but abnormal X-rays) and 152 were considered not to have TB (not TB). Oral swabs RT-PCR were positive in 29/80 (36.3%) bacteriologically-confirmed, 9/34 (26.5%) bacteriologically-negative and 29/152 (19.1%) not TB. The yield varied among samples stored for less and more than 30 days (p = 0.013) from 61% (11/18) and 29% (18/62) among bacteriologically confirmed, and 30.8% (4/13) and 23.8% (5/21) among bacteriologically-negative participants. Among not TB patients, the specificity was 80.9% (123/152), being 78.3% (18/23) among samples stored less than 30 days and 81.4% (105/129) among samples stored for more than 30 days (p = 0.46). The detection of M. tuberculosis in oral mucosa samples is feasible, but storage conditions may affect the yield.

scholarly journals Cases Report of Cured Novel Coronavirus-infected Pneumonia Patients with Viral Nucleic Acid Test Positive in Fecal Specimens

2020 ◽  
Author(s):  
Mei Han ◽  
Jingbo Zou ◽  
Wenguang Tian ◽  
Xiaoyu Wei ◽  
Yang Zhou ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Molecular Diagnostic ◽  
Clinical Practices ◽  
Rt Pcr ◽  
Viral Nucleic Acid ◽  
Nucleic Acid Test ◽  
Assessment Standard ◽  
Novel Coronavirus ◽  
Acid Test ◽  
Respiratory Specimens

Abstract Background: The outbreak of the novel coronavirus in China (COVID-19) represents a significant and urgent threat to global health. We report here five cases of COVID-19 infection patients in our clinical practices who are medically stable and presumed to successfully “cleared” the virus after antiviral treatments. Case presentation: The clinical evaluation depends on the viral nucleic acid test in respiratory specimens by real-time PCR reverse transcription (RT-PCR) assays according to the authorized guidance. We found that the stool samples of these cured patients remain positive in RT-PCR assay while the virus is undetectable in respiratory specimens. RT-PCR molecular diagnostic assay was designed to specifically detect the presence of viral RNA. Thus, the positive result in the fecal specimens implies the existence of viable virions with the patients. Conclusions: This highlights the importance to look closely at the assessment standard of medical treatment, as well as the need for reevaluation of the criteria for the initial screening, prevention, and care of patients with this emerging infection.

scholarly journals Detection and Localization by In Situ Molecular Biology Techniques and Immunohistochemistry of Hepatitis C Virus in Livers of Chronically Infected Patients

1998 ◽  
Vol 46 (5) ◽  
pp. 653-660 ◽  
Author(s):  
Francine M. Walker ◽  
Marie-Christine Dazza ◽  
Marie-Christine Dauge ◽  
Olivier Boucher ◽  
Christophe Bedel ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Interferon Α ◽  
Rt Pcr ◽  
Biopsy Specimens ◽  
Bile Ductule ◽  
Liver Biopsies ◽  
Cytoplasmic Signals ◽  
Detection And Localization

Hepatitis C virus (HCV) detection in the livers of chronically infected patients remains a debatable issue. We used immunohistochemistry, in situ hybridization (ISH) alone or after microwave heating with FITC-labeled probes, RT-PCR with unlabeled primers followed by ISH (RT-PCR-ISH), and in situ RT-PCR with FITC-labeled primers (in situ RT-PCRd) to localize the virus in 38 liver biopsy specimens from 21 chronically infected HCV patients treated with interferon-α (IFN-α). Biopsies were taken at the beginning and end of IFN-α treatment and 1 year later. Results were compared with that of HCV-PCR in serum. RT-PCR-ISH and in situ RT-PCRd showed HCV signal in all liver biopsies even in responders with seronegative HCV PCR. This signal was intranuclear, diffuse, or peripheral, in hepatocytes, bile ductule cells, and lymphocytes. Cytoplasmic signals were occasionally observed. Whereas the percentage of labeled hepatocytes remained constant, the number of labeled lymphoid follicles decreased after INF-α therapy. Immunohistochemistry resulted in the same pattern of positivity but it was weaker and inconstant. This study indicates the persistency of HCV latency in IFN-α responders 1 year after IFN-α treatment cessation, a finding that certainly deserves confirmation.

Recent advances in the laboratory diagnosis of peste des petits ruminants

2021 ◽  
Vol 77 (05) ◽  
pp. 226-231
Author(s):  
WIESŁAW NIEDBALSKI ◽  
ANDRZEJ FITZNER ◽  
KRZYSZTOF BULENGER ◽  
ANDRZEJ KĘSY
Keyword(s):  
Reverse Transcription ◽  
Animal Health ◽  
Clinical Signs ◽  
Small Ruminants ◽  
Clinical Diagnostics ◽  
Molecular Diagnostic ◽  
Molecular Diagnostic Techniques ◽  
Peste Des Petits Ruminants ◽  
Rt Pcr ◽  
Agar Gel

Peste des petits ruminants (PPR) is a highly contagious and economically important, viral disease of small ruminants caused by the peste des petits ruminants virus (PPRV), which belongs to the genus Morbilivirus in the family Paramyxoviridae. PPR control is achieved mostly through vaccination and/or slaughter of susceptible animals coupled with clinical or laboratory-based diagnosis. Since clinical signs of PPR are not disease-specific and clinical diagnostics is not reliable, it should be confirmed by laboratory testing. Laboratory confirmation of clinical suspicions is made by detection of PPRV in blood, swabs or post-mortem tissues through classical virus isolation (VI), agar gel immunodiffusion (AGID)/agar gel precipitation test (AGPT), counter-immunoelectrophoresis (CIE), immunoperoxidase test (IPT) or enzyme-linked immunosorbent (ELISA) assays. However, these conventional methods have been superseded by more rapid, sensitive and accurate molecular diagnostic techniques based on the amplification of parts of either nucleocapsid (N) or fusion (F) protein gene, such as RT-PCR, real-time RT-PCR, reverse transcription loop-mediated isothermal amplification (RT-LAMP), reverse transcription recombinase polymerase amplification (RT-RPA) and Oxford nanopore MinION technology. Although these molecular diagnostic assays are accurate, rapid and sensitive, they have to be performed in laboratory settings, and samples must be transported under appropriate conditions from the field to the laboratory, which can delay the confirmation of PPRV infection. The recently developed immunochromatographic lateral flow device (IC-LFD) assay can be used in the field (“pen-side”) without the need for expensive equipment, so a well-established laboratory is not required. The control and eventual eradication of PPR is now one of the top priorities for the Food and Agriculture Organization (FAO) and the World Organization for Animal Health (OIE). In 2015, the international community agreed on a global strategy for PPR eradication, setting 2030 as a target date for elimination of the disease

Expression and intracellular localization of Nanos2-homologue protein in primordial germ cells and spermatogonial stem cells

Zygote ◽  
2019 ◽  
Vol 27 (02) ◽  
pp. 82-88 ◽  
Author(s):  
Vivek Pandey ◽  
Anima Tripathi ◽  
Pawan K. Dubey
Keyword(s):  
Stem Cells ◽  
Flow Cytometry ◽  
Germ Cells ◽  
Expression Patterns ◽  
Intracellular Localization ◽  
Primordial Germ Cells ◽  
Type A ◽  
Spermatogonial Stem Cells ◽  
Single Type ◽  
Rt Pcr

SummaryThe decision by germ cells to differentiate and undergo either oogenesis or spermatogenesis takes place during embryonic development and Nanos plays an important role in this process. The present study was designed to investigate the expression patterns in rat of Nanos2-homologue protein in primordial germ cells (PGCs) over different embryonic developmental days as well as in spermatogonial stem cells (SSCs). Embryos from three different embryonic days (E8.5, E10.5, E11.5) and SSCs were isolated and used to detect Nanos2-homologue protein using immunocytochemistry, western blotting, reverse transcription polymerase chain reaction (RT-PCR) and flow cytometry. Interestingly, Nanos2 expression was detected in PGCs at day E11.5 onwards and up to colonization of PGCs in the genital ridge of fetal gonads. No Nanos2 expression was found in PGCs during early embryonic days (E8.5 and 10.5). Furthermore, immunohistochemical and immunofluorescence data revealed that Nanos2 expression was restricted within a subpopulation of undifferentiated spermatogonia (As, single type A SSCs and Apr, paired type A SSCs). The same results were confirmed by our western blot and RT-PCR data, as Nanos2 protein and transcripts were detected only in PGCs from day E11.5 and in undifferentiated spermatogonia (As and Apr). Furthermore, Nanos2-positive cells were also immunodetected and sorted using flow cytometry from the THY1-positive SSCs population, and this strengthened the idea that these cells are stem cells. Our findings suggested that stage-specific expression of Nanos2 occurred on different embryonic developmental days, while during the postnatal period Nanos2 expression is restricted to As and Apr SSCs.

scholarly journals Varicocele Repair Improves Testicular Histology in Men with Nonobstructive Azoospermia

2015 ◽  
Vol 2015 ◽  
pp. 1-5 ◽  
Author(s):  
Murat Ustuner ◽  
Hasan Yilmaz ◽  
Ufuk Yavuz ◽  
Seyfettin Ciftci ◽  
Ali Saribacak ◽  
...  
Keyword(s):  
Extraction Procedure ◽  
Testicular Biopsy ◽  
Testicular Tissue ◽  
High Serum ◽  
Histopathological Analysis ◽  
Nonobstructive Azoospermia ◽  
Biopsy Specimens ◽  
Testicular Histology ◽  
Late Maturation ◽  
Varicocele Repair

Objective. To determine the histopathological differences after varicocele repair in testicular tissue in males with nonobstructive azoospermia.Methods. Between 2009 and 2014, 45 men with complete azoospermia and palpable varicocele, presenting with primary infertility of at least 1 year, undergoing varicocele repair at our institution were selected for the study. A standard systematic testicular 6-core Tru-Cut biopsy was performed during varicocele repair. Other biopsies were obtained from each testicle of all patients at the time of microscopic sperm extraction procedure.Results. Nineteen patients were selected for the study. Testicular biopsy specimens were classified as Sertoli cell only on preoperative histopathological analysis in 14 patients. After varicocele repair, focal spermatogenesisn=3and late maturation arrestn=2were found in these patients. Average Johnsen score was significantly increased after varicocelectomyP=0.003. Motile sperm was found in one patient on postoperative semen analyses and in 10 more patients in the microscopic sperm extraction procedure. Preoperative high serum follicle stimulating hormone level and venous reflux were significantly and negatively correlated with the increase in average Johnsen scoreP<0.05.Conclusions. Our findings suggest significant improvement in testicular histology after varicocele repair.

Classical and Molecular Cytogenetic Abnormalities in 124 Pediatric Patients with Acute Lymphocyte Leukemia.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4419-4419
Author(s):  
Yihuan Chai ◽  
Hui Lv ◽  
Jun Lu ◽  
Peifang Xiao ◽  
Jianqing Li
Keyword(s):  
Bone Marrow ◽  
Multiplex Pcr ◽  
Cytogenetic Analysis ◽  
Chromosomal Abnormalities ◽  
Molecular Diagnostic ◽  
Pcr Analysis ◽  
Rt Pcr ◽  
Childhood All ◽  
Cytogenetic Abnormalities ◽  
Conventional Cytogenetic Analysis

Abstract In childhood acute lymphocyte leukemia (ALL), cytogenetics plays an important role in diagnosis, allocation of treatment and prognosis. On base of the conventional cytogenetic analysis, molecular methods have inproved our ability to accurately and rapidly risk-stratify patient with childhood ALL in the last few years. Our aim was to assess the demography of cytogenetic abnormalities in childhood ALL. The study sample consisted of 124 newly diagnosed ALL patients younger than 16 years, who were diagnosed at the Department of Pediatric Hematology/Oncology, Soochow University Children’s Hospital. The diagnosis and FAB subtypes of ALL was determined by Wright-Giemsa-stained bone marrow smears and cytochemicalstaining. Immunophenotyping of the bone marrow samples was performed by flow cytometry. Multiplex reverse transcription-polymerase chain reaction (RT-PCR) analysis was performed to detect the 29 most common leukemia translocations for routine molecular diagnostic hematopathology practice, and complement the information gained from conventional cytogenetic analysis. Cytogenetic analysis was successful in 112 of 124 children with ALL. Sixty-eight (60%)of them had clonal chromosomal abnormalities. Numerical imbalances consisted of hyperdipoild(>47 chromosomes, 36 cases), hypodipoild(<46 chromosomes, 14 cases), pseudodiploidy(18 cases). Chromosomal translocations were observed in 13 patients by conventional cytogenetic analysis. Three cases were found positive for t (4;11), 3 cases for t (9;22), 1 case for t (1;19) and 6 cases for other rare translocations. RT-PCR analysis detected 116 of the 124 ALL patients. Thirteen cases of TEL-AML1, 10 cases of rearrangement in the MLL gene, 4 cases of E2A-PBX1, 4 cases of E2A-HLF, 3 cases of BCR-ABL, 2 cases of TLS-ERG, 32 cases of HOX11, were detected by RT-PCR in B-lineage leukemias. SIL-TAL1 had been found in 4 of 7 of T-lineage leukemias. Sixty-eight cases of ALL show chromosomal aberrations. Multiplex PCR positivity was detected in 59(50%)of the 116 ALL patients studied. Multiplex PCR combined with chromosome analysis uncovered Chromosomal abnormalities in 95 of 124(77%) of ALL patients and supplemented each other in detecting Chromosomal abnormalities. It provides reliable evidence for the diagnosis, classification and prognosis.

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